System for automated compression of chromatography columns

ABSTRACT

The present invention provides an automated system and method for maintaining compaction, and therefore increased efficiency, of a media bed within a chromatography column. In the preferred embodiment, an adjustment assembly is slidingly engaged inside one end of the column such that it can be moved along the column&#39;s major axis. When idle, the force exerted on this end is equal to the compression on the media. When the column is actively processing chromatographic fluid, this exerted force can be expressed as the sum of the compression on the media, and the force of the fluid being processed. This total force and the fluid pressure are monitored using a load cell and a pressure sensor respectively. The compression force operating on the media bed is then computed based on these measurements and compared to the optimal value. The position of the adjustment assembly within the column is then modified in response to changes in the measured compression force.

This application is a divisional application of U.S. patent applicationSer. No. 11/604,458, which in turn was a divisional application of U.S.patent application Ser. No. 11/072,081, the disclosures of which areincorporated herein by reference.

BACKGROUND OF THE INVENTION

The present invention relates to chromatography columns and inparticular to a chromatography column system and method of compressingand maintaining optimal or a consistent compression on a media bedwithin a chromatography column. Frequently it is desirable to separateout one or more useful components from a fluid mixture that containsother components that may not be useful or are less valuable. Toaccomplish this, it is often necessary or desirable to fractionate sucha fluid mixture to separate out the useful or desired components. Thiscan be carried out by using liquid chromatography systems. Liquidchromatography may be described as the fractionation of components of amixture based on differences in the physical or chemical characteristicsof the components. The various liquid chromatographic systemsfractionate the components with a fractionation matrix. Some liquidchromatographic matrix systems fractionate the components of a mixturebased upon such physical parameters as molecular weight. Still otherliquid chromatographic systems will fractionate the components of amixture based upon such chemical criteria as ionic charge,hydrophobicity, and the presence of certain chemical moieties such asantigenic determinants or lectin-binding sites on the components.

Chromatography systems of various sizes are used in both laboratoryanalysis operations and for industrial scale production operations inwhich separation steps such as separating out a fraction from humanblood or separating out impurities from a pharmaceutical can be carriedout on a large scale in a batch process.

Separations using chromatography columns filled with chromatographicmedia have been carried out for years. The chromatographic mediatypically comprises particles having a diameter between 5 and 100 μm. Tomaximize the effectiveness of the column, it is desirous to arrange themedia as tightly and as uniformly as possible. This process, known aspacking, eliminates voids and channels within the media. However,chromatography column packing, particularly where large columns areinvolved, is highly variable and can dramatically affect the efficiencyof the separation. Many setup process parameters must be smoothlyorchestrated in order to achieve a homogenous packed column. Dependingon the size of the column, the packing process can take a significantamount of time, in the range of several hours. Yet despite the timeinvested in packing the column, often times less than 50% of thesepacked columns function in accordance with the specification.

During chromatography packing and operation, the compaction of thechromatographic media has a significant impact on the performance andrepeatability of the column. In packing the column, typically the mediais compressed through an alternating process of flowing liquid throughthe column to pack the media and then lowering the adjuster assembly inan effort to mechanically compress the media.

Once the column has been packed, the fluid to be fractionated is thenpassed through the column. During extended operation, packed media bedswill experience a variety of issues with the media.

In some media, as it is wet with the packing and/or process fluid, itswells. This can cause an overcompression of the media potentiallydamaging the media or leading to a decrease in separation efficiency dueto a reduction in the media pore sizes or availability of the media tothe process stream.

Also a slight, but noticeable and cumulative bed compaction occurs. Thisis intrinsic to many packed bed columns. The sources of this furthercompaction are principally process-dependent and are generally due tothe hydraulic drag of processing, flow perturbations during processcycles, mobile phase properties, such as flow rate, viscosity anddensity, support matrix swelling, and the intrinsic bead mobility in thepacked bed superstructure. The magnitude of the compaction is alsorelated to the size, shape and rigidity of the medium particles. Forexample, irregularly shaped particles such as PROSEP® matrix will bepredisposed to de-bridging and the accompanying compaction.

The long-term result is that as a packed bed is repeatedly used, itincrementally compacts. This compaction leads to a continuous reductionin media bed density at the top of the bed, until a breach forms betweenthe bed and the top bed support. The immediate performance implicationsare a decrease in separation efficiency, typically characterized bybroader elution peaks with more tailing. Ultimately, this reduced beddensity can lead to the formation of preferential flow channels throughthe bed, decreasing the effective life of the column, therebynecessitating more frequent repacking.

Therefore, there is a need for an improved method of maintaining propercompression within a packed chromatography column during operation,which will improve the performance of the column and extend its usefullife.

SUMMARY OF THE INVENTION

The problems of the prior art have been overcome by the presentinvention, which provides an automated system and method for maintainingcompaction, and therefore increased efficiency, of a media bed within achromatography column. In the preferred embodiment, an adjustmentassembly is slidingly engaged inside one end of the column such that itcan be moved along the column's major axis. When idle, the force exertedon this end is equal to the compression of the media bed. When thecolumn is actively processing, this exerted force can be expressed asthe sum of the compression of the media, and the force of the fluidbeing processed. This total force and the fluid pressure are monitoredusing a load cell and a pressure sensor, respectively. The compressionforce operating on the media bed is then computed based on thesemeasurements and compared to the desired or preferably optimal value.The position of the adjustment assembly within the column is thenmodified in response to changes in the measured compression force tomaintain a consistent compression on the media bed.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a cross sectional view of a first embodiment of the presentinvention.

FIG. 2 is a cross sectional view of a second embodiment of the presentinvention.

FIG. 3 is a cross sectional view of the preferred embodiment of thepresent invention.

FIG. 4 shows the embodiment of FIG. 3 in the pivoted position.

DETAILED DESCRIPTION OF THE INVENTION

The repeated processing of a chromatography column generally willtypically cause the further compaction of the previously packed media.This compaction can be significant. For example, in one trial, usingmedia, repeated separation of a milieu of E. Coli proteins wasperformed. During the first separation process, the bed height wasmeasured to be 56 cm. After the fortieth separation, the bed height hadbeen compacted to a height of only 48 cm. This compaction led to abreach between the top surface of the media bed and the top bed supportand decreased efficiency.

There is also a second issue associated with the bed height. The supportmatrices of the resins used in the media can change in volume. Eachmatrix possesses its own swelling behavior, with dextran-based andcellulosic resins being most susceptible to swelling when subjected topH changes. Ionic strength also has a significant impact on the swellingof cellulosic, agarosic and dextran-based chromatography media such asion exchangers. Generally, this swelling is most pronounced during theelution, regeneration, and most particularly the cleaning phases of achromatographic separation cycle. Therefore, the column must be capableof adapting to swelling-induced changes in the media bed to preventover-pressurization of the column, or overstressing of the media.

FIG. 3 illustrates the preferred embodiment of the present invention.Before use, chromatography column 110 is filled with media slurry in amanner known to those skilled in the art. The adjustable bed support112, which forms a tight seal along the walls of column 110, is thenmoved down inside the column tube 110. Typically, the adjustable bedsupport has a cross-sectional configuration that matches that of thecolumn. Preferably, the bed support also has a gasket, or other sealingmeans along its perimeter to ensure the tightness of the seal. Thisallows the buffer within the column 110 to flow out the bottom flow port113. Generally one bed support is fixed in place while the other is freeto move. Alternatively, both supports can move if desired. In theembodiment as shown the bottom bed support 114 is fixed in place to thecolumn. During this packing process, a media bed 120 forms and iscontacted by the adjustable bed support as it continues to apply forceto the bed 120. Thus, when the bed 120 is fully compacted, it exerts aforce on adjustable bed support 112.

Adjustable bed support 112 is coupled to a shaft 130, which ispreferably threaded. Shaft 130 passes through an opening 141 in yoke140, which opening is also preferably threaded. Yoke 140 is held inposition by stanchions 150, which are mounted to a base 160, on whichthe column 110 preferably rests. In the preferred embodiment, thestanchions 150 are held in contact with the base through the use offasteners 161, such as bolts, which extend through openings 164 in thebase and engage with the stanchion via slots 151 bored into thestanchion, which are also threaded. The fastener has a shaft 162, whichis preferably threaded, of a given diameter, and a head 163 having adiameter larger than that of the shaft. The openings 164 in the base 160are preferably larger than the diameter of the fastener's shaft 162, butsmaller than the diameter of the fastener's head 163, to allow thefastener's shaft to move freely through the opening 164. The fastener161 is inserted from the underside of the base 160, through the opening164 such that the fastener's shaft 162 engages with the slot 151 in thestanchion 150.

Yoke 140 is affixed to a plurality of stanchions 150. Two stanchionstypically provide the needed structural stability for smaller diametercolumns, while additional stanchions may be used for large diametercolumns. These stanchions 150 are preferably placed equidistant from oneanother around the circumference of a circle that is concentric to, butlarger than column 110. The stanchions 150 have a height equal to, orpreferably greater than, that of the column 110.

In one embodiment, yoke 140 is connected to the two or more stanchionsand it spans the width and centerline of the column 110. The yoke 140 isretained to the stanchions 150 by means such as slot 152, a ring orother device that can affirmatively hold the yoke 140 in place. The yoke40 may be permanently attached to the stanchions 150 or more preferably,it may be removably connected to the stanchions 150 by bolts, clevispins, cotter pins, clamps and the like. In one preferred embodiment, theyoke 40 is attached to one stanchion 50 by a bolt, and the otherstanchion by a clevis pin so that when adjustable bed support 112 iswithdrawn from the column, the yoke 140 can be pivoted vertically aboutstanchion 150 containing the bolt and moved up and out of the way of thecolumn to allow easy access to the column interior. FIG. 4 shows thatembodiment in the pivoted position.

In another embodiment, the yoke 140 can also rotate in a horizontalcircular motion away from the mouth of the column 110.

Atop the yoke 140 is an actuator 170 adapted to move the shaft in thevertical direction, independent of the yoke 140. This actuator can bepneumatically, electrically or hydraulically controlled. In thepreferred embodiment, a motor, preferably electrically powered, isequipped with a gear that contacts the threaded shaft 130. The movementof the motor causes the rotation of the gear, which in turn causesrotation of the threaded shaft 130. The resulting rotation of thethreaded shaft 130, through the threaded opening 141 in yoke 140 causesthe shaft 130 to move relative to the yoke 140 in the verticaldirection.

The adjustable bed support 112, shaft 130, and actuator 170 comprise theadjuster assembly. These components operate in unison to adjust theposition of the adjustable bed support 112 inside the column 110,thereby also controlling the pressure exerted on the media bed.

The yoke 140 and the stanchions 150 comprise a support structure 155.This structure is rigidly coupled and is affixed to the shaft 130 andthe base 160, such that any force exerted on adjustable bed support 112is transferred through shaft 130, through support structure 155, to theconnection point between the support structure 155 and the base 160.

While this embodiment comprises a preferred embodiment in which a singleshaft with 2 stanchions is used, the invention is not so limited. Thoseskilled in the art will appreciate that it is within the scope of thepresent invention to use multiple shafts and a greater number ofstanchions. For example, a very large diameter column may require agreater number of shafts and stanchions in order to insure that theadjustable bed support descends uniformly and evenly onto the media bed.

Alternatively, other structures can be utilized. Chromatography columnsare formed of three basic components; a column tube, a bottom fixed endand a top, movable end. See U.S. Pat. No. 4,350,595 and U.S. Pat. No.6,139,732. The top end moves relative to the tube so as to be capable ofremoval for introduction and removal of chromatography media in the tubeand to be capable of longitudinal travel into the tube to compress themedia for use.

This top end however needs to be fixed at some point to the column inorder to move relative to the column.

A first means for accomplishing this is to form a tube of high strengthmaterials, including metals such as stainless steel or rigid structuralplastics, such as acrylics or polymethylpentenes such as TPX® plasticavailable from Mitsui Petrochemical Industries Ltd Corporation of Japan.The tube has a flange at the upper end to which a top plate is attachedto the column and a flange at the lower end to which a fixed bottom endis attached. The top, movable end is then attached to this top plate andtravels relative to it in and out of the tube.

In one embodiment, shown in FIG. 1, the tube 2 has a bottom plate 4fixed in place by bolts 6 attached to a flange 8 of the tube 2. A topplate 10 is fixed to a top flange 12 of the tube 2 by setscrews 13. Amovable end 14 is centrally located in the top plate 10 and is capable,by movement of rod 16, of moving into or out of the tube 2.

As the end 14 moves into the tube 2 to compress the media bed 18 foruse, longitudinal forces are carried from the end 14 to the rod 16 tothe top plate 10 and then to the tube 2 itself.

Another alternative is shown in FIG. 2. It uses a series of rods 20 orscrews closely aligned around the outside of the tube 24 to carry thelongitudinal forces rather than the wall of the tube itself. This allowsone to use less structurally rigid materials, such as glass or plastics,preferably acrylic or styrene, and to also use thinner walled tubes. Allof this reduces the weight and cost of the device.

Most of the elements of that tube 24 of FIG. 2 are similar to those ofFIG. 1. One has a movable top end plate 22, a bottom plate 26, attachedto a fixed bottom end 27, flanges 28, either as part of the tube 24 orin this example as separate pieces to secure the fixed top plate 30 andbottom plate 26 to the tube 22. A rod 32 extends through the plate 30and is connected to the movable end 22 by a handle 34. A bed ofchromatography media 36 is compressed by the movement of the end 22.Also shown in FIG. 2 are a series of guide rods 38, which are used, inlarger columns to keep the end 22 horizontal during movement. Plate 30is normally affixed on flange 33 and attached by numerous mechanicalfasteners 31.

In the preferred embodiment, a load cell 180 is located between the head163 of the fastening device and the underside of base 160. However, theload cell 180 can be positioned in any location where it can measure theforce exerted on the media bed. For example, the load cell can bepositioned between bolt 6 and bottom plate 4 in FIG. 1. Similarly, theload cell can be located between mechanical fastener 31 and plate 30 orbetween mechanical fastener 31 and bottom plate 26 in FIG. 2. A loadcell is a device that translates the load exerted on it into an analogelectrical output, such as voltage or current. The relationship betweenthe exerted load and the electrical output is well established andtightly controlled, such that the exact load experienced by the loadcell can be determined by monitoring its electrical output. The termload cell is used herein to include any device that carries out thisfunction.

Returning to FIG. 3, the load cell 180 is preferably circular, with aconcentric opening in the middle, such that the diameter of the openingis large enough to allow shaft 162 to be slid through the opening.However, the diameter of the opening is preferably smaller than thediameter of the head 163 of the fastener, such that the head cannot passthrough the opening, thereby causing the load cell to interconnect withthe fastener in a similar manner as a traditional washer. Thus, thefastener is inserted through the concentric opening in the load cell180, through the opening in the base 160, and into the slot of stanchion150. Preferably, one load cell is used, regardless of the number ofstanchions, however multiple load cells, or one load cell for eachstanchion, are also envisioned as an embodiment of the presentinvention.

One skilled in the art will appreciate that although the preferredembodiment comprises an adjustable top bed support, and a fixed lowerbed support, the invention is not so limited. The apparatus can also beconstructed such that the top support is fixed, and the lower bedsupport is adjustable.

In the preferred embodiment, the fluid to be processed by the column 110travels in a conduit through a hollow cavity within shaft 130 toadjustable bed support 112. Alternatively, the fluid may also travel ina conduit parallel to the shaft and then enter the adjustable bedsupport under a hollow arch formed at the base of the shaft. Adjustablebed support 112 also comprises a flow cell, which equally distributesthe fluid such that it enters the media bed uniformly. The processedfluid then exits the column through bottom flow port 113. Those skilledin the art will appreciate that the direction of the fluid's travel isnot limited to top to bottom; the fluid can also be forced into thebottom of the column and drawn out of the top surface. Similarly, it isnot required that the fluid entry and the movable support be located inthe same end of the column.

The pressure of the fluid entering the column is monitored. There are anumber of methods known in the art for performing this monitoring. Forexample, a bubble trap can be inserted between the source of the fluidand the entrance to the shaft 130. A pressure sensor associated with thebubble trap can be used to supply the measured fluid pressure. In thepreferred embodiment, a pressure sensor 190, preferably a transducer, isin communication with the fluid flow through the use of a T connectionin close proximity to the shaft 130. A pressure transducer is used toconvert a pressure measurement into either an analog or digitalelectrical signal, such as voltage or current. In this scenario, thetransducer 190 measures the pressure of the fluid being forced throughthe conduit and into the column 110.

Having defined the components of the present invention, the operationnow will be described. First, the media in the column is compressed toform a media bed. This process can be accomplished in a variety of wayswell known to those skilled in the art, and the present invention is notlimited to a specific packing methodology. Once the column has beenpacked, the adjustable bed support 112 will be in direct contact withthe top of the bed 120, holding it under some amount of sufficient forceto insure that it remains compacted. The media bed 120 exerts acounterforce onto the adjustable bed support 112. Since the adjustablebed support 112 is rigidly affixed to the shaft 130, which is rigidlyaffixed to the yoke 140, which is in turn rigidly affixed to thestanchions 150, this exerted force is transferred directly to thefastener 161 which is securing the stanchion 150 to the base 160. Thus,the force exerted by the media bed 120 is measurable by load cell 180,located between fastener 161 and the underside of the base 160. In thepreferred embodiment, a single load cell is utilized, thus this loadcell will experience only a fraction of the total force exerted by themedia bed. That fraction is defined as 1/(# of stanchions). Thus, if twostanchions are utilized, the load cell will experience of the totalforce exerted by the media bed 120. Alternatively, load cells can beplaced in association with each stanchion. In this case, the total forcewould be defined as the sum of the forces experienced by each load cell.Similarly, if load cells are arranged on only a portion of thestanchions, the total load can be expressed as:

(Σ of load cells)*(# of stanchions)/(# of load cells).

The outputs from the pressure sensor 190 and the load cell 180 are incommunication with controller 100. Controller 100 also generates outputsto the actuator 170 directing it to alter the position of the adjustablebed support. By using the output from the load cell 180 in conjunctionwith controller 100, it is then possible to create a control system,whereby the load experienced by the load cell is used by the controller100 to adjust the position of the shaft 130, using actuator 170. Oneskilled in the art will appreciate that the controller can be of varioustypes, including, but not limited to proportional,proportional-derivative (PD), proportional integral (PI) orproportional-integral-derivative (PID), and that the invention is notlimited by the choice of the controller. Similarly, the output from thecontroller 100 to the actuator 170 can be in various forms, includingbut not limited to analog voltage, current, digital signals, or pulses.

The optimal force to be applied to the media bed 120 can be determinedusing a number of different methods, such as but not limited toempirical measurements as the column is packed, or fixed values based onthe amount and type of media being used. Once the optimal force requiredto create the proper compression on the media bed 120 is determined, thecontrol system comprising the load cell 180, actuator 170 and controller100 operate to maintain this force. The method of determining thisoptimal force is independent of the present invention, and therefore anymethod of determining this value is suitable.

Having established the proper compression for the media bed 120, thecolumn is then ready to accept fluid. The fluid that enters the columnwill also be under pressure, and this pressure will also be exerted onthe adjustable bed support. Therefore, the total force exerted on theadjustable bed support can be given by:

F _(Total) =F _(Fluid) +F _(Media Compression),

and F_(Fluid) is given by:

F _(Fluid) =P _(Fluid) *Area _(Adjustable Bed Support).

Combining these, the total force on the adjustable bed support is:

F _(Total) =P _(Fluid) *Area _(Adjustable Bed Support) +F_(Media Compression).

Since the total force can be measured via the load cell, and the fluidpressure can be measured via the pressure sensor 190, it is possible todetermine the amount of force being applied to the media bed.

F _(Media Compression) =F _(Total) −P_(Fluid)*Area_(Adjustable Bed Support).

This computed F_(Media Compression) is then compared to the optimalcompression force. By adjusting the position of shaft 130 based on themeasurement from the load cell 180 and the pressure sensor 170, it ispossible to maintain a constant optimal pressure on the media bed 120.For example, as the media bed compresses, it will exert less force onthe adjustable bed support 112. This reduction will be measured by theload cell as a decrease in total force (assuming a constant fluidpressure). The controller will detect this reduced force, and willdetermine that the force being exerted on the media bed has decreased.To compensate for this, the controller will actuate the actuator 170 toadjust the shaft 130 to further compress the column, until the mediacompression force returns to the optimal value. Conversely, if the mediabed expands, the controller detects an increase in total force and willactuate the actuator 170 to retract the shaft 130 in a direction out ofthe column, until the media compression force returns to the optimalvalue.

The control system of the present invention can be utilized in a numberof different ways. In a first embodiment, the control system is usedonly between separation cycles to correct for any changes in the heightof the media bed 120 that occurred during the previous cycle. In thisembodiment, the position of the adjustable bed support within thecylinder is held constant throughout the separation cycle, and then itsheight is adjusted after the completion of the cycle.

In a second embodiment, the control system is continuously operational,thereby constantly adjusting the pressure exerted on the media bed bychanging the position of the adjustable bed support within the cylinder.Due to the precision required, this embodiment preferably utilizes a PIDcontroller. One skilled in the art will recognize that a continuouscontrol system can be approximated through the use of a sampled system,whereby the load cell and pressure transducers are sampled at periodicintervals and adjustments to the vertical position of the adjustable bedsupport are made in response to these sampled measurements.

1. The process of setting the compression force on a media bed in achromatography system, said system comprising a column having an inletand containing a media bed through which a fluid is adapted to flow; anadjustable bed support movable in said column and adapted to exert aforce on said media bed; a sensor for determining the pressure of saidfluid entering said column; a load cell for measuring the force exertedon said adjustable bed support; and an actuator for moving saidadjustable bed support in response to the force measured by said loadcell, said process comprising the steps of: using said load cell tomeasure said force exerted on said adjustable bed support; calculatingcompression force on said media bed, based on measurement from saidload; comparing said calculated compression force to an optimalcompression force; and using said actuator to move said adjustable bedsupport based on said comparison.